HNMT

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Abstract

Betaine homocysteine methyltransferase (BHMT) is a Zn²⁺-dependent thiolmethyltransferase that regulates homocysteine levels, the elevation of which is considered a risk factor for cardiovascular disease. Most plasma homocysteine is produced through the liver methionine cycle, where BHMT metabolizes approximately 25% of this non-protein amino acid. This process allows recovery of one of the three methylated equivalents used in phosphatidylcholine synthesis by methylation. BHMT has been known for over 40 years, but the difficulties encountered in its isolation have only recently been studied in detail.

Introduction

In recent years, homocysteine (Hcy) has attracted much attention due to its role as a risk factor for cardiovascular disease. The increases in plasma levels (tHcy) have also been detected in disorders such as psoriasis, renal insufficiency, spina bifida, Alzheimer's disease, cognitive impairment in the elderly, and adverse pregnancy outcomes. In fact, elevated tHcy levels may be detected in 10-20% of the normal population due to both genetic and non-genetic factors. The liver processes approximately 48% of the ingested methionine in humans, so it has been proposed that the liver is the main regulator of tHcy levels. Hcy, a non-protein thiol amino acid is generated through a reaction catalyzed by S-adenosylhomocysteine hydrolase (SAHH) that breaks the S-adenosylhomocysteine (SAH) produced through S-adenosylmethionine (SAM)-dependent methylations into Hcy and adenosine. The reaction is reversible and thermodynamically favors the synthesis of SAH, a potent inhibitor of many methylation reactions. In order to control the level of this inhibitor, the elimination of reaction products must be coordinated with their synthesis.

Figure 1. Homocysteine metabolism in the liver (Pajares, M. A.; Pérez-Sala, D. 2006)

The BHMT gene

The BHMT gene contains 8 exons and 7 introns and is located on chromosomes 5q13.1-5q15 in humans. A BlastN search revealed 61% identity between human and mouse genes. Analysis of the 5' end of genomic DNA revealed a possible TATA box located 26 bp and 28 bp upstream of the transcription initiation site of the human and mouse genes, respectively. In addition, 5′ of the human TATA box putative sites for several transcription factors are located, including Sp1 (four sites), activator protein 2 (one site), liver-specific or liver-enriched factors HNF-1, HNF-3 and CAAT-enhancer-binding protein, homeobox 4c, 4d and 4e and steroid receptors. This area of the mouse gene contains putative Sp1, AP2, AP4, NF1, GATA and CCAAT sites. Two additional Sp1 sites have been found in human genes at 3' of the TATA box. Some negative regulatory elements are located in the region comprising –1793/–1063 with compensatory elements are located in –3175/–1793. No apparent effect was observed with the addition of methionine or ethionine to the HepG2 medium, whereas SAM reduced activity. The SAM response element is at –254/+1. However, these experimental data only identify putative regulatory sites, so more in-depth studies of the role of these regulatory elements are warranted.

Figure 2. BHMT structure (Pajares, M. A.; Pérez-Sala, D. 2006)

Interactions of BHMT

The extraordinary abundance of BHMT in the liver and its presence in the lens, which does not produce betaine, both point to its potential effects in processes other than methionine synthesis. To date, interacting partners including apoB mRNA, tubulin, hepatitis B virus (HBV), and transglutaminase have been identified by screening cDNA expression and yeast two-hybrid libraries, 2D gel electrophoresis, and immunoprecipitation. In addition, BHMT2 and BHMT fragments were found in a prion protein (PrP) screening and in autolysosomal and lysosomal membranes, respectively, binding to microtubules was observed by colocalization using confocal microscopy, and coassembly of BHMT with tubulin dimers was also obtained. In addition, transglutaminases modify BHMT at its C-terminus, or generate cross-linked 260- and 520-kDa multimers. Because the fragments lacking part of the N-terminus may adopt different folding patterns, and therefore function differently. This new fold may also result from binding to tubulin or post-translational modifications.